Field of the Invention
This invention broadly relates to affinity chromatography wherein the specific recognition of one chemical for a second is used to identify the presence of either. One such application is found in the immunoassay of immunochemicals such as antigens and antibodies. This invention is accordingly discussed in the content of immunoassay apparatus and, more particularly, in the context of apparatus including an immunochemically coated member and a receptical tray having a fluid-containing well sized to hold both a fluid sample and the coated member.
As used herein, the term "immunochemical" is generic to antigens and antibodies. For the purpose of this disclosure, a select immunochemical is defined as one whose presence or absence is to be determined, while an immobilized immunochemical is one which reacts with the select chemical to permit identification of the select chemical. Accordingly, when the select immunochemical is an antibody, the immobilized immunochemical will be an antigen specific thereto and, conversely, a select antigen will require the use of an immobilized antibody. For the sake of clarity, further discussion will be limited to a typical immunoassay process wherein the immobilized chemical is an antigen, the select chemical is an antibody and the fluid sample is one of human blood. It is to be understood, however, that other permutations and applications are within the scope of the invention.
Several direct and indirect methods for detecting the select immunochemical are known in the art. An example of a commonly practiced direct method employs competitive binding. A primary surface containing the immobilized immunochemical (typically an antibody to a hormone) is placed in a sample containing a known amount of labelled select immunochemical (typically, labelled hormone). Any (unlabelled) select immunochemical present in the sample competes with the labelled immunochemical for attachment to the immobilized immunochemical. Accordingly, a decrease in the latter from that obtained when a control sample containing only the labelled immunochemical is used, may be used to quantify the concentration of the former.
In the indirect method the coated member is coated with the immobilized immunochemical, such as the antigen of a particular virus, and immersed in the sample, where select antibodies, if any, become attached to the antigens. The member is removed, washed in an aqueous buffer solution, which is compatible with the antigen, to remove nonspecific reactions and placed in a second solution having a labelled reagent which recognizes the select immunochemical. The member is washed a second time to remove unbound labelled reagents and assayed.
Two labelling methods commonly used are radioisotopic means wherein the radioactively tagged reagent is detected by means of a counter, and enzyme-tagged means wherein the addition of a substrate to the tagged reagent produces a color change indicative of select immunochemical concentration. As will be apparent, the present invention is applicable to both direct and indirect methods as well as to both the radioisotopic and enzyme-tagged labelling methods.